A clinical test is routinely carried out which involves converting a component to be measured in a biological sample, such as serum, into hydrogen peroxide using an oxidase and measuring the generated hydrogen peroxide to determine the component to be measured. There is often a problem of the influence of interfering substances, such as ascorbic acid and bilirubin, contained in a biological sample, in a method for measuring a component to be measured in the biological sample based on the measurement of hydrogen peroxide. Particularly, ascorbic acid has an extremely strong reduction effect; to avoid the influence thereof, a method involving converting ascorbic acid to dehydroascorbic acid using an ascorbic acid oxidase to suppress the influence of ascorbic acid is often used in a clinical test.
Ascorbic acid oxidase is a blue copper protein having a molecular weight of about 140,000 and converts ascorbic acid to dehydroascorbic acid. To suppress the influence of ascorbic acid, an ascorbic acid oxidase is often contained in a reagent for measuring a component to be measured in a biological sample. However, there is a problem that an ascorbic acid oxidase is unstable and is deactivated during the preservation of the reagent to deteriorate the performance of the reagent.
Against the problem, there are known a method for stabilizing an ascorbic acid oxidase by adding a compound having both of a metal or a positive basic group and a negative acid group, or a combination of the compound, catalase and/or peroxidase, and an oxidative dye coupler to a solution containing an ascorbic acid oxidase (see patent document 1); a method for stabilizing an ascorbic acid oxidase by maintaining an ascorbic acid oxidase under lyophilization in the presence of an N-substituted taurine buffer (see patent document 2); a method for stabilizing an ascorbic acid oxidase by allowing an ascorbic acid oxidase to coexist with one or more selected from skim milk, lactose, sucrose, and soluble starch (see patent document 3); a method for stabilizing an ascorbic acid oxidase in a solution state by chemically binding an ascorbic acid oxidase to a water-soluble carrier selected from bovine serum albumin, dextran, and polyethylene glycol (see patent document 4); a method for stabilizing an ascorbic acid oxidase by adding one or more kind of pyruvic acid (and salts thereof) therein (see patent document 5); a method for stabilizing an ascorbic acid oxidase by adding a sugar alcohol to a solution containing an ascorbic acid oxidase (see patent document 6); a method for stabilizing an ascorbic acid oxidase by allowing the ascorbic acid oxidase to coexist with a substance selected from 2-methyl-4-isothiazolin-3-one, 5-chloro-2-methyl-4-isothiazolin-3-one, 1,2-benzisothiazolin-3-one, and salts thereof (see patent document 7); a method for stabilizing an ascorbic acid oxidase in a dried state by allowing the ascorbic acid oxidase to coexist with a protein degradation product (see patent document 8); a method for stabilizing an ascorbic acid oxidase by adding an amino acid and an amino acid salt and/or an oligopeptide (see patent document 9), and the like.